Tuberculosis (TB) is a complex disease and a major cause of mortality world-wide. Despite the development of new treatments, TB remains a global health concern. Annually, there are more than seven million new cases and two million deaths caused by TB (C. Martin, Eur Respir J 2005, 26:162-167). Coinfection with HIV leads to an exacerbation of the disease and contributes to higher mortality in HIV patients. Programs to combat TB in many countries have failed to eradicate TB, partly due to the spread of multidrug-resistant TB and the low efficacy of the BCG vaccine. Therefore, the exploration of novel drug targets and vaccines against Mycobacterium tuberculosis (Mtb), the main causative pathogen of TB, is essential.
Among pathogenic bacteria, Mtb causes more deaths in humans than any other pathogen. Approximately one third of the world population has already been infected by Mtb. Mtb is an intracellular pathogen that has evolved to persist efficiently in infected macrophages. The composition of the Mtb cell wall is important for the interaction with host cells during the initial steps of the infection. Later, cell wall components play a crucial role in modulating the pro-inflammatory response by macrophages and also serve as a protective barrier to prevent anti-tuberculosis agents from permeating inside. Consequently, the antibiotics used for the treatment of tuberculosis require long term administration (J. C. Sacchettini, E. J. Rubin and J. S. Freundlich, Nat Rev Microbiol 2008, 6:41-52). Mortality in people living in developing countries is high since their access to these antibiotics is often limited and compliance with treatment courses is low.
The major components of the mycobacterial cell wall are the mycoyl arabinogalactan-peptidoglycan (mAGP) complex and interspersed glycolipids including ManLAM, LAM, LM, and PIMs. While the mAGP complex is covalently attached to the bacterial plasma membrane, the glycolipids are non-covalently attached through their phosphatidyl-myo-inositol (PI) anchor. PIMs constitute the only conserved substructure of LM, LAM and ManLAM (FIG. 1). The inositol residue of PI is mannosylated at the C-2 position to form PIM1 and further at the C-6 position to form PIM2, one of the two most abundant naturally occurring PIMs, along with PIM6. Further α-1,6 mannosylations give rise to PIM3 and PIM4, the common biosynthetic precursors for PIM5, PIM6 and the much larger LM structures. LAM is constituted by attachment of arabinans—the repeating units of α-1,5 arabinose terminated with a single β-1,2 arabinose—to unknown mannose units of LM. The non-reducing end arabinose in the arabinan moiety of LAM can be capped at the C-5 position with one or two α-mannose units to furnish ManLAM.
Among the surface components involved in the Mtb interaction with host cells, PIMs play a crucial role in the modulation of the host immune response. The functional importance of PIMs was emphasized by the finding that PIMs bind to receptors on both phagocytic (J. B. Torrelles et al., J. Immunol. 2006, 177:1805-1816) and nonphagocytic (H. C. Hoppe et al., Infect Immun 1997, 65:3896-3905) mammalian cells. Recently, it has been shown that PIMs, but neither LAM nor ManLAM interact with the VLA-5 on CD4+T lymphocytes and induce the activation of this integrin (R. E. Rojas et al., J Immunol 2006, 177:2959-2968). These'findings suggest that PIMs are not only secreted to the extracellular environment, but also exposed on the surface of Mtb to interact with host cells.
Although different functions have been ascribed to the PIMs, it remains to be determined whether and to which extent the different PIM substructures display biological activity. A better understanding of the mycobacterial cell wall biosynthesis is required to be able to counteract with the problems of drug resistance and bacterial persistence. Synthetic PIMs represent important biochemical tools to elucidate biosynthetic pathways and to reveal interactions with receptors on host cells. PIMs are potential vaccine antigens and/or adjuvants.
Several synthetic PIMs containing fewer mannoside units have been synthesized employing various chemical methodologies. In contrast to PIM3 and PIM4 that contain only α-1,6 mannosidic linkages, PIM5 and PIM6 also incorporate α-1,2 mannosides that might contribute to different biological activities of these PIMs. None of the studies to date utilized synthetic PIMs that contain linkers for immobilization. Coupling of synthetic PIMs to carrier proteins, beads, quantum dots, microarray or surface plasmon resonance (SPR) surfaces opens a host of options for biochemical studies.